This invention relates to nucleic acid and amino acid sequences of ATP synthase and to the use of these sequences in the diagnosis, prevention, and treatment of cancer, myopathies, neurodegenerative diseases, and diseases and disorders of the immune and sympathetic nervous systems.
The mitochondrial electron transport (or respiratory) chain is a series of enzyme complexes in the mitochondrial membrane that is responsible for the transport of electrons from NADH to oxygen and the coupling of this oxidation to the synthesis of ATP (oxidative phosphorylation). ATP then provides the primary source of energy for driving a cell""s many energy-requiring reactions.
ATP synthase (F0F1ATPase) is the enzyme complex at the terminus of this chain and serves as a reversible coupling device that interconverts the energies of an electrochemical proton gradient across the mitochondrial membrane into either the synthesis or hydrolysis of ATP. This gradient is produced by other enzymes of the respiratory chain in the course of electron transport from NADH to oxygen. When the cell""s energy demands are high, electron transport from NADH to oxygen generates an electrochemical gradient across the mitochondrial membrane. Proton translocation from the outer to the inner side of the membrane drives the synthesis of ATP. Under conditions of low energy requirements and when there is an excess of ATP present, this electrochemical gradient is reversed and ATP synthase hydrolyzes ATP. The energy of hydrolysis is used to pump protons out of the mitochondrial matrix.
ATP synthase is, therefore, a dual complex, the V portion of which is a transmembrane proton carrier or pump, and the F1 portion of which is catalytic and synthesizes or hydrolyzes ATP. Mammalian ATP synthase complex consists of sixteen different polypeptides (Walker, J. E. and Collinson, T. R. (1994) FEBS Lett.346: 39-43). Six of these polypeptides (subunits xcex1, xcex2, xcex3, xcex4, xcex5, and an ATPase inhibitor protein, IF1) comprise the globular catalytic F1ATPase portion of the complex, which lies outside of the mitochondrial membrane. The remaining ten polypeptides (subunits a, b, c, d, e, f, g, F6, OSCP, and A6L) comprise the proton-translocating, membrane spanning F0 portion of the complex. Like other members of the respiratory chain, all but two of the polypeptide subunits of ATP synthase are nuclear gene products that are imported into the mitochondria; a and A6L are products of mitochondrial genes. Enzyme complexes similar to mammalian ATP synthase are found in all cell types and in chloroplast and bacterial membranes. This universality indicates the central importance of this enzyme to ATP metabolism.
Transcriptional regulation of these nuclear encoded genes appears to be the predominant means for controlling the biogenesis of ATP synthase. Defects and altered expression of ATP synthase and other enzymes in the respiratory chain are associated with a variety of disease conditions in man, including neurodegenerative diseases, myopathies, and cancer.
The discovery of polynucleotides encoding ATP synthase, and the molecules themselves, provides a means to investigate the control of cellular respiration under normal and disease conditions. Such molecules related to ATP synthase satisfy a need in the art by providing new diagnostic or therapeutic compositions useful in diagnosing and treating cancer, myopathies, neurodegenerative diseases, and diseases and disorders of the immune and sympathetic nervous systems.
The present invention features two ATP synthase subunits, designated individually as Asy-1 and Asy-2 and collectively as Asy, and characterized as having similarity to the ATP synthase subunits.
Accordingly, the invention features substantially purified Asy proteins Asy-1 and Asy-2, having the amino acid sequences shown in SEQ ID NO:1 and SEQ ID NO:3, respectively.
One aspect of the invention features isolated and substantially purified polynucleotides that encode Asy proteins-Asy-1, Asy-2. In a particular aspect, the polynucleotides are the nucleotide sequences of SEQ ID NO:2 and SEQ ID NO:4, respectively.
The invention also features a polynucleotide sequence comprising the complement of SEQ ID NO:2, or SEQ ID NO:4, or variants thereof. In addition, the invention features polynucleotide sequences which hybridize under stringent conditions to SEQ ID NO:2 or SEQ ID NO:4.
The invention additionally features nucleic acid sequences encoding polypeptides, oligonucleotides, peptide nucleic acids (PNA), fragments, portions or antisense molecules thereof, and expression vectors and host cells comprising polynucleotides that encode Asy. The present invention also features antibodies which bind specifically to Asy, and pharmaceutical compositions comprising substantially purified Asy. The invention also features the use of agonists and antagonists of Asy.